The broad objective of this proposal is to enable embryonic stem cells (ESCs) to specifically recognize and bind to infarcted tissues, which they are expected to replace and regenerate. Cardiac repair by stem cell implantation holds promise as a post-infarct therapeutic option. Compared to intra- myocardial injection, intra-vascular infusion of stem cells is less invasive and better tolerated by patients with acute myocardial infarction (MI). However, the relatively low rate of ESC retention in the infarct site after transvascular infusion is a limiting factor that prevents sufficient engraftment and regeneration. To address the challenge, we will modify the stem cell surface so that transfused cells are able to bind to infarcted tissue by recognizing a well-defined molecular marker of cell death, anionic phospholipids (APLDs). APLDs become exposed to the extracellular milieu during apoptosis and necrosis, thereby providing a specific and ample binding target for the recognition of infarcted tissue by cognate moieties. Such a cognate moiety is the C2A domain of synaptotagmin I, a well- characterized APLDs-binding protein that recognizes dead and dying cells with high specificity and affinity. Recently, we demonstrated that intravenous-injected C2A avidly accumulates in infarcted myocardium. To logically extend this intriguing finding, we will apply this unique molecular recognition mechanism to mediate the engraftment of intravascularly delivered ESCs into infracted myocardium. The primary goal is to confer infarct avidity to ESCs by attaching the C2A to the ESC surface. As part of the Preliminary Studies, we have demonstrated that ESCs can be surface-labeled with C2A, without affecting cellular physiology or pluripotency. And, as a proof of feasibility, we have observed that C2A-conjugated ESCs bind dead cardiomyocytes, but not viable cardiomyocytes. In light of these preliminary data, we hypothesize that ESCs equipped with APLDs-binding activity will lead to a significantly improved engraftment within infarcted sites in vivo. To enhance therapeutic cardiac repair and to test this hypothesis, we propose the following Specific Aims: 1) to refine and optimize the conjugation methods for attaching the C2A to the surface of ESCs;and 2) to assess the engraftment rate of C2A-conjugated ESCs in infarcted cardiac tissue and evaluate the long-term cardiac function in a mouse model of acute MI. Once established, the concept of conferring infarct avidity should be applicable to other types of transplanted cells, including differentiated cell types derived from ESCs. Overall, this work explores a novel transplantation strategy that will aid in cardiac repair with translational potential. PUBLIC HEALTH RELEVANCE: Stem cell therapy holds promise in regenerating infarcted myocardium. While the intra-vascular delivery of stem cells is minimally invasive and better tolerated by patients, its efficacy is limited by a low retention rate of infused cells in the infarct site. The goal of this proposal is to make stem cells capable of recognizing and binding to infarcted tissues. This will be accomplished by attaching an infarct-avid protein to the stem cell surface without altering cellular physiology. Making stem cells with a binding affinity toward dead and dying tissues will not only help target transplanted stem cells to the infarct site, but also improve their rate of retention and engraftment within the infarcted tissues.